Equine laminitis, which is sometimes referred to as foundering, is a common disorder that has been recognized and described in even the earliest books of veterinary medicine. In an article by Green, M. E. et al., which appeared in Equine Medicine and Surgery, Fourth Edition, Vol. II, Colahan, P. T. et al. (Eds.), American Veterinary Publications, Inc. (1991), Chapter 12, pp. 1354-1358, laminitis is described as an inflammation of the pedal laminae that form the supportive bond between the hoof and the third phalanx. Laminitis is further described as a disorder of the hoof whose cause is varied. It is a complex, multi-systemic disease affecting the digestive, cardiovascular, hemic, renal, endocrine, musculoskeletal, integumentary, and immune systems. It is characterized by multi-systemic aberrations that ultimately result in reduced capillary perfusion, ischemia, and necrosis of the laminae. These results are accompanied by pain and loss of supportive function. Acute laminitis is described as comprising the events leading up to and the onset of lameness. Acute laminitis can progress to the chronic stage. The chronic stage ensues after persistent lameness (greater than 48 hours), or when the distal phalanx deviates detectably. Chronic laminitis is a consequence of some degree of loss of integrity of the supporting digital laminae. A photograph of a horse suffering from the effects of laminitis of the forelimbs is shown on FIG. 1. Notice that the horse assumes a recumbent position in which most of the weight is placed on the hind limbs and little weight bearing pressure is exerted on the forelimbs.
There are reports of evidence that equine laminitis is caused by ingestion of too much grain; colic; retained placenta; exhaustion; ingestion of black walnut shavings; ingestion of too rich grass; excessive concussion, and/or excessive cold water. Despite the gathering of voluminous information over a period exceeding 300 years, equine laminitis still remains incompletely understood, however.
While it is possible that an equine that has been subjected to this condition can recover without intervention treatment, during the course of the disease there is substantial pain, recumbency, hoof wall deformation and even sloughing of the hooves. Several symptoms are displayed, including: a bounding digital pulse, warm feet, an abnormal gait, a shifting of weight, or some combination of some or all of these. If the animal does not recover, after the onset of lameness, the laminae deteriorate, the animal's feet are extremely painful, and the coffin bone becomes displaced. Destruction of the animal is then the only humane course of action.
Because of the serious consequences of this condition, it has been the subject of many and varied treatments over the years. In almost all cases, since the condition appears to be a function of a loss of circulation in the hoof area, the treatments have been directed to increasing the blood supply to and circulation within the hoof and adjacent tissues.
There are several different treatments that are currently being used to care for horses with laminitis. Several different vasodilators have been used in the past to treat this disorder and some are still in use. Many of them involve the use of nitroglycerin, which has been applied transdermally. Other methods use orally administered isoxsuprine, also a known vasodilator. Current therapy also includes the use of anticoagulants, such as heparin, aspirin and trental (pentoxifylline). All current therapies suffer from one or more drawbacks, including difficulty in the mode of administration of the active drug, lack of effectiveness, lack of compliance and lack of simplicity in the proposed treatment regimen.
Katsuki, S. et al., in J. Cyclic Nucleotide Res. (1977) 3:23-35, point out that the activity of a variety of smooth muscle relaxing agents, including sodium nitroprusside, nitroglycerin and sodium nitrite, may be related to their ability to increase tissue levels of cyclic guanosine monophosphate (cGMP) or to the formation of nitric oxide (NO). Other nitrogen-containing compounds may also function similarly, including hydroxylamine or sodium azide. These substances may also possess vasodilatory properties. These and other nitrogen-containing compounds may then be referred to as nitric oxide "donors" or "precursors."
It has been shown that vascular endothelial cells synthesize nitric oxide from L-arginine (L-Arg) but not D-arginine. See, Palmer R. M. J. et al., in Nature (1988) 333:664-666. This article also describes how the release of NO from endothelial cells induced by bradykinin and the calcium ionophore A23187 is reversibly enhanced by infusions of L-Arg or L-citrulline. The release of NO by certain cells can be protracted. Hence, L-Arg can be considered an NO donor or precursor, but because the conversion of L-Arg may not immediate and may occur over a protracted period of time, L-Arg can be thought of as a "slow acting" NO precursor. As used herein a "slow acting" NO precursor may provide for the release of NO in the tissues over the course of a few hours to several hours after initial exposure to the "slow acting" NO precursor. In contrast, a "fast acting" NO precursor, such as nitroglycerin or nitroprusside, generally provides almost instantaneous release of detectable levels of NO in the plasma or tissue (e.g., within about a few minutes of exposure to the "fast acting" NO precursor and lasting for several minutes). Such "fast acting" agents may be quickly depleted, however. Thus, a fast acting NO donor may be used to provide a burst of NO, while a slow acting NO donor may be used to provide a more sustained, protracted level of NO release.
An abstract by Hinckley, K. A. et al., which appeared in J. Endocrinol. (1994) 143:P103, illustrates the complexity of the aetiology of laminitis. These authors conclude that the causes of laminitis are multifactorial. These authors speak only of the intravenous administration of L-Arg.
In an editorial leader in the publication, Equine Vet. J. (1996) 28(1):1-2, Elliott, J. discusses the merits of nitric oxide treatment of laminitis. In this article, it is pointed out that NO was formerly known as endothelium derived relaxing factor, a tribute to the fact that this gas is produced continuously by the lining of blood vessels as a result of the action of an enzyme present in the endothelial cells, endothelial nitric oxide synthase or eNOS. It is thought that eNOS is activated by a rise in intracellular calcium concentration, catalyzing the conversion of L-Arg to NO and L-citrulline. The nitric oxide passes through biological membranes, binds to haem iron in the soluble enzyme guanylate cyclase (GC). The activity of GC is thus stimulated, cyclic guanosine monophosphate concentrations increase and vascular smooth muscle tone is reduced. There is speculation in this article that disruption of the blood flow to the sensitive laminae of the equine foot, which occurs in laminitis, might involve some disturbance in the L-Arg-NO pathway.
The subject of the editorial, an article by Hinckley et al. in the same issue, Equine Vet. J. (1996) 28(1):17-28, describes a treatment that uses intravenously administered L-Arg and topically applied nitroglycerin (glyceryl trinitrate or GTN). The L-Arg is administered i.v. as a 10% aqueous solution, while the GTN is provided as a 20% ointment through a patch that is positioned, using adhesive tape, over digital vessels. In particular, a laminitic pony, weighing 250 kg, received a total dose of 120 g of L-Arg or 0.48 g/kg bwt at a rate of approximately 40 mL/min (16 mg/kg bwt/min) for 30 min. The GTN patches were also applied once daily to three limbs only, 12 hours after the L-Arg infusion. Drops in heart rates were observed, including hypotension and cardiac arrhythmia, with the i.v. infusion. Some ponies showed signs of pain, sweating and shivering. The topical application of GTN patches appeared to improve the lameness of some of the treated ponies. No improvement was observed in others. Thus, it is clear that L-Arg, given intravenously, is not a particularly effective treatment. The results of the balance of this study are also mixed. There is no suggestion in this article that L-Arg should be administered in a manner other than intravenously or that any benefits may be obtained from a further pursuit of a combination treatment.
Lecithin organogels are obtained by adding a minimal amount of water to a solution of lecithin in organic solvent. Typically, a minimum amount of water is added to lecithin dissolved in an organic solvent. See, Scartazzini, R. and Luisi, P. L., in J. Phys. Chem. (1988) 92:829-833. The results of an investigation on the transdermal transport of scopolamine and broxaterol through human skin using a soybean lecithin organogel has been described. Willimann, H. et al., in J. Pharmaceutical Sci. (1992) 81(9):871-874. It was found that the rate of transdermal transport of scopolamine was about one order of magnitude higher with the organogel relative to an aqueous solution of the drug at the same concentration. However, the rate of transport of the drug was not different with the organogel compared with a pre-gelation, microemulsion of the drug (lecithin, organic solvent and drug, but no added water).
In U.S. Pat. No. 4,882,164, granted to Ferro, A. and Steffen, H., an aqueous mixed micelle solution composed of cholanic acid salts and lipids is used for the solubilization of non-steroidal anti-inflammatory drugs (NSAIDs). Cholanic acid salts include cholates, glycocholates and taurocholates. Lipids include phosphatidylcholines, especially, natural and synthetic lecithins. A formulation in accordance with the invention is described, as follows: carprofen (50 mg), L-Arg (30 mg), anhydrous glycocholic acid (88.5 mg), 40% sodium hydroxide (19 microliters), lecithin for mixed micelles (169 mg), 2N HCl (to pH 6.0) and water for injection (to make 100 mL final volume). The presence of the cholanic acid is essential to the composition described. Further, the presence of an NO precursor or vasodilator is not disclosed, taught, or suggested.
NSAIDs are compounds that are structurally different from steroids and which display an anti-inflammatory activity. NSAIDs usually contain a carboxylic acid group and are derived from acetic acid, propionic acid, butyric acid, salicylic acid and the like. Subcategories of NSAIDS, include aminoarylcarboxylic acids (e.g., enfenamine, flufenaminic acid, isonixin, niflumic acid and the like), arylacetic acids (e.g., acemetacin, alcoflenac, amfenac, 30 clopirac, felbinac, fenclofenac, ibufenac, indomethacin, isoxepac, sulindac, zomepirac and the like), arylbutyric acids (e.g., bumadizon, fenbufen, xenbucin and the like), arylcarboxylic acids (e.g., clinadac, ketorolac and the like), arylpropionic acids (e.g., benoxaprofen, carprofen, ibuprofen, ibuproxam, indoprofen, ketoprofen, naproxen, oxaprozin, protizinic acid, suprofen and the like), pyrazoles, pyrazolones and thiazinecarboxamides. Specific examples of NSAIDs include, but are not limited to, flunixin, epirizole, apazone, feprazone, ramifenazone, droxicam, aspirin, phenylbutazone, piroxicam, bendazac, ditazol, guaiazulene, oxaceprol, proxazole, tenidap and the like, or the salts thereof.
Hence, a strong need remains for an effective treatment of equine laminitis.